Process for the isolation of galanthamine

ABSTRACT

The subject matter of the present invention concerns a process for the isolation of the alkaloid galanthamine; the galanthamine itself which has been produced according to this process; the use of the galanthamine thus produced in galenic preparations; and the galanthamine thus produced for treating narrow-angle glaucoma, Alzheimer&#39;s disease, as well as alcohol and nicotine dependence.

1. The present invention relates to a process for isolating the alkaloidgalanthamnine; the galanthamine itself which has been produced by thisprocess; the use of galanthamine thus manufactured in galenicpreparations; as well as to galanthamine thus produced for the treatmentof narrow-angle glaucoma, Alzheimer's disease, as well as alcohol andnicotine dependence.

2. Galanthamine (4a, 5, 9, 10, 11, 12-hexahydro-3-methoxy-11-methyl-6H-benzofuro-(3a, 3, 2-ef) - (2) benzazepine-6-ol) is a tetracyclicalkaloid which, owing to its pharmacological properties, belongs to thegroup of reversibly acting cholinesterase inhibitors. Its effects aresimilar to those of physostigmine and neostigmine. However, it also hasown specific properties, for example, highly analgesic effectscomparable to those of morphine. As a cholinesterase inhibitor thetherapeutic range of galanthamine is three to six times broader, sinceit is not as toxic as physostigmine and neostigmine. This advantagecompensates for its slightly lower cholinesterase-inhibiting action,relative to dosage. Galanthamine is used in poliomyelitis and differentdiseases of the nervous system, but mainly in the treatment ofnarrow-angle glaucoma and as antidote after curare applications. The useof galanthamine in the treatment of Alzheimer's disease is in anexperimental stage. Lately, the treatment of alcohol and nicotinedependence has also been described (DE-OS 40 10 079, DE-OS 43 01 782).

3. The therapy of Alzheimer's disease, alcohol and nicotine dependence,as well as that of narrow-angle glaucoma requires long-acting drug formsadapted to the particular circumstances. This could be an eye ointmentin the treatment of narrow-angle glaucoma. In the treatment ofAlzheimer's disease, alcohol or nicotine dependence, complicatedtherapeutic plans or prolonged infusions are unsuitable for obviousreasons. In these diseases, a transdermal therapeutic system (TTS) is asuitable form, for example that described in DE-OS 43 01 783. Neitherthe intact skin nor the cornea permit absorption of active substancesalts. For this reason it is not possible to use galanthaminehydrobromide or galanthamine hydrochloride with ointments or a TTS inthe therapy of narrow-angle glaucoma, Alzheimer's disease, or alcohol ornicotine dependence. The pure galanthamine base must therefore be used.

4. An economically efficient synthesis of galanthamine base is notpossible because of its complicated tetracyclic structure with threeoptically active carbon atoms. For this reason, galanthamine is usuallyisolated from plants belonging to Amaryllidaceae, for example, fromgalanthus species, such as the snowdrop or Leucojum aestivum. Theseplants have the advantage of comprising galanthamine in concentrationsof up to 0.3% with only small amounts of companion alkaloids so that theextraction method described in DE-PS 11 93 061 can be used. However,both the galanthus species and Leucojum aestivum are protected. On theother hand, the extraction method described in DE-PS 11 93 061preferably uses chlorohydrocarbons which have fallen into discredit fortoxicological reasons. The Pharmacopoeias of the Western World thereforecall for a limitation of the residual chlorohydrocarbon content to <10ppm. In the preparation of drugs chlorohydrocarbons should therefore beavoided as far as possible. Moreover, in the known process, the solventextract must be adsorbed to alumina to ensure separation of the resinoussubstances and companion alkaloids. From the solution obtained afterfiltering off the alumina, the galanthamine is then purified bygalanthamine hydrobromide; this involves disposal of halogen salts. Forthe use in ointments or in a TTS, the galanthamine base must thenadditionally be liberated from this galanthamine hydrobromide.

5. It is accordingly the object of the present invention to provide aprocess for the isolation and purification of galanthamine, which doesnot have the drawbacks of the prior art processes. In particular,purification is to be facilitated, the use of chlorohydrocarbons andpurification by means of galanthamine salts are to be avoided. Theobject is achieved according to the present invention by a processhaving the characterizing Features of claim 1. Preferred embodiments arecharacterized in the subclaims.

6. In detail, the subject matter of the present invention is a processfor the isolation of galanthamine from biological material which isrecovered from agriculturally cultivated amaryllidacea species or fromthose which are commonly regarded as “weeds” and are not protected,preferably from the bulbs of these plants. These amaryllidaceae include,for example, narcissi or crinum species. Particularly suitable areNarcissus pseudonarcissus “Carlton” or the Asian climber Crinum amabile.Although these plants only have a tenth of the galanthamine amountcontained in the protected plants, and, moreover, have up to twelvecompanion alkaloids, the process according to the present inventionsurprisingly makes it possible to isolate galanthamine base therefromwith a purity suitable for the use in drugs.

7. According to the present invention galanthamine-containing,biological material, which is preferably communicated and mixed withalkali powder, preferably with sodium hydroxide pellets, soda, potash,or similar salts suitable to liberate bases from biological material andto prepare pharmacological active substances, is extracted withtoxicologically safe organic solvents, and the galanthamine from thisextract is purified by means of liquid-liquid extraction. It isparticularly important for the success of the process according to thepresent invention to maintain the pH-value in the first stage of theliquid-liquid extraction. The liquid-liquid extraction is carried out ina first stage at a pH of about 4, and in a second stage at a pH of about9. In addition to concentrated ammonia, soda solution of another basemay be used to adjust the pH-value. Diethyl ether or specialboiling-point gasoline is used as toxicologically safe organic solvent.Special boiling-point gasoline is preferably used, with it a certainpreliminary purification can be achieved. The solvent is removed;galanthamine is recrystallized from a suitable solvent, preferably fromisopropanol. The white galanthamine base having a melting point of 129to 130° C. is obtained. The purity of the galanthamine thus obtained isshown in the HPLC-chromatogram (FIG. 2).

8. This result is very amazing since the process according to DE-PS 1193 061 used so far was found to be unsuitable to isolate galanthaminefrom biological material containing small amounts of galanthamine and upto 12 companion alkaloids. The process described in DE-PS 11 93 061results in an emulsion which cannot be broken so that an extraction isrendered impossible. A slightly modified process resulted in an oilyresidue which, according to HPLC-chromatogram (FIG. 1), is contaminatedby at least four substances.

9. It must therefore be regarded as a particular surprise that the freegalanthamine base can be isolated in pure form and good yield frombiological material comprising small amounts of galanthamine in additionto a large number of companion alkaloids by means of the simple processaccording to the present invention, without having to carry out anyadditional adsorption step to aluminum oxide.

10. The galanthamine base produced according to the present inventionmay be used for the treatment of narrow-angle glaucoma, Alzheimer'sdisease, or alcohol and nicotine dependence. In addition, thegalanthamine base produced according to the present invention may beused in galenic preparations, such as ophthalmic ointments ortransdermal therapeutic systems, which may also be used to treat theabove-mentioned diseases.

11. The present invention is more particularly described in the examplesthat follow, which are intended to illustrate, but not to limit, thepresent invention:

EXAMPLE 1

12. (Comparison):

13. In accordance with the process described in DE-PS 11 93 061, 10 kgair-dried, communicated bulbs of Narcissus pseudonarcissus “Carlton” iscarefully mixed with 2.5 l of a 8% aqueous ammonia solution. Thematerial swells; the whole batch pastes up. The addition of 23 ldichloroethane intended for extraction results in an emulsion whichcannot be b.,token.

EXAMPLE 2

14. (Comparison):

15. The process for the isolation of galanthamine known in the art(DE-PS 11 93 061) is modified. 10 kg air-dried, communicated bulbs ofNarcissus pseudonarcissus “Carlton” is carefully mixed with 400 g sodiumcarbonate. 23 l dichloroethane is added. The mixture is allowed to standfor 10 hours; then the solvent is decanted. The bulbs are once againdoused with 23 l dichloroethane which is decanted after 2 to 3 hours.After that, 17 l dichloroethane is added to the bulks for the thirdtime; however, this is decanted immediately. The mixed dichloroethaneextracts are extracted by means of 10% sulfuric acid (2×600 ml each;2×300 ml each). The acidic extracts are mixed and purified from tracesof dichloroethane by means of shaking out with diethyl ether. Understirring and cooling to 15 to 20° C., about 200 ml of a 25% aqueousammonia solution is then added up to alkaline litmus reaction. The pH isin the range of 7 to 8. Different from the indications in the art, thecompanion alkaloids do not precipitate. The alkaline solution issaturated with salt and extracted with diethyl ether. After evaporationof the ether, a negligible residue remains, which is also different fromthe indications in the art. The pH-value of the aqueous phase is set toabout 14 by saturating it with potash. The aqueous phase is repeatedlyextracted with diethyl ether. The mixed ether extracts are evaporated todryness, the remaining galanthamine-containing residue is dissolved inacetone (50 ml). In contrast to the art, there is no precipitate. 350 mlacetone is replenished, 200 g aluminum oxide is added, and stirring iseffected for 45 minutes. The aluminum oxide is filtered off and washedtwice with 100 ml acetone each time. The mixed acetone solutions areevaporated to dryness. 1.3 g of an oily residue is obtained which isexamined by means of HPLC. The chromatogram is shown in FIG. 1. The mainpeak, the galanthamine, is marked. It is clearly visible that thegalanthamine thus isolated is contaminated by at least four substances.

EXAMPLE 3

16. 100 kg air-dried, communicated bulbs of Narcissus pseudonarcissus“Carlton” is carefully mixed with 4 kg of sodium carbonate. The mixtureis divided into three equal parts, and each is doused with 15 l specialboiling-point gasoline 80/110. The mixtures are allowed to stand for 24hours. The solvents are each renewed twice, collected, and evaporated todryness in low vacuum. The extracts are placed in 2% aqueous sulfuricacid and adjusted to a pH of 4 with concentrated aqueous ammoniasolution. Five extractions with diethyl ether follow. The aqueous phaseis set to a pH of 9 with concentrated ammonia and extracted five timeswith diethyl ether. These ether fractions are collected, dried withsodium sulfate, and evaporated. 20 g of a slightly yellow, oily residueis obtained, which is recrystallized from hot isopropanol. 10 g of whitegalanthamine base having a melting point of 129-130° C. is obtained.Only one pealing is visible in the HPLC chromatogram (FIG. 2).

1. A process for the isolation of galanthamine characterized in thatgalanthamine-containing, biological material is extracted withtoxicologically safe organic solvents and that the galanthamine fromthis extract is purified by means of liquid-liquid extraction, saidliquid-liquid extraction being carried out in the first stage at apH-value of about
 4. 2. The process according to claim 1 characterizedin that the biological material is communicated and mixed with alkalipowder prior to the extraction.
 3. The process according to any one ofclaims 1 or 2 characterized in that the liquid-liquid extraction incarried out in the second stage at a pH-value of about
 9. 4. The processaccording to any one of claims 1 to 3 characterized in that sodiumcarbonate is used as alkali powder.
 5. The process according to any oneof claims 1 to 4 characterized in that diethyl ether or specialboiling-point gasoline is used as toxicologically safe organic solvent,special boiling-point gasoline being preferred.
 6. The process accordingto any one of claims 1 to 5 characterized in that the obtainedgalanthamine is recrystallized from a suitable solvent.
 7. The processaccording to claim 6 characterized in that Isopropanol is used assolvent.
 8. The process according parts of amaryllidaceae, are used asbiological material.
 9. The process according to claim 8 characterizedin that bulbs are used as plant parts.
 10. The process according to anyone of claims 1 to 9 characterized in that narcissi species or crinumspecies are used as amaryllidaceae.
 11. The process according to claim10 characterized in that as narcissi species narcicissus pseudonarcissus“Carlton”, or as crinum species Crinum amabile are used. 12.Galanthamine in pharmaceutical quality with a purity of ≧99%, producedaccording to a process in accordance with any one of claims 1 to 11 .13. Galenic administration forms characterized by a content ofgalanthamine in accordance with claim 12 .
 14. The use of thegalanthiamine according to claim 12 for the production of ophthalmicointments.
 15. The use of the galanthamine according to claim 12 for theproduction of transdermal therapeutic systems.
 16. The use of thegalanthamine according to claim 12 for the production of a drug for thetreatment of narrow-angle glaucoma.
 17. The use of the galanthamineaccording to claim 12 for the production of a drug for the treatment ofAlzheimer's disease.
 18. The use of the galanthamine according to claim12 for the production of a drug for the treatment of alcohol dependence.19. The use of the galanthamine according to claim 12 for the productionof a drug for the treatment of nicotine dependence.
 20. A pharmaceuticalpreparation characterized in that it comprises galanthamine according toclaim 12 in addition to pharmaceutical adjuvants known per se.
 21. Thepharmaceutical preparation according to claim 20 for topicalapplication.
 22. The pharmaceutical preparation according to claim 20 or21 for the treatment of narrow-angle glaucoma.
 23. The pharmaceuticalpreparation according to claim 20 or 21 for the treatment of Alzheimer'sdisease.
 24. The pharmaceutical preparation according to claim 20 or 21for the treatment of alcohol dependence.
 25. The pharmaceuticalpreparation according to claim 20 or 21 for the treatment of nicotinedependence.
 26. A process for the therapeutic treatment of the humanorganism characterized in that galanthamine according to claim 12 or apharmaceutical preparation according to any one of claims 20 or 21 isused.